Nutraceutical compositions and use thereof

ABSTRACT

Compositions including, as the active ingredients, resveratrol, a derivative, metabolite or analogue thereof, and at least one additional component selected from epigallocatechin gallate (EGCG), genestein, vitamin E, polyunsaturated fatty acids, gamma-linolenic acid and vitamin K are disclosed. Also disclosed, are methods of making such compositions including admixing resveratrol, a derivative, metabolite or analogue thereof, and at least one additional component selected from EGCG, genestein, vitamin E, polyunsaturated fatty acids, gamma-linolenic acid and vitamin K are disclosed. Methods for the treatment or prevention of inflammatory diseases including administering to a subject in need of such treatment resveratrol, a derivative, metabolite or analogue thereof, in combination with at least one additional component selected from epigallocatechin gallate (EGCG), genestein, vitamin E, polyunsaturated fatty acids, gamma-linolenic acid and vitamin K are also disclosed.

CROSS-REFERENCE TO RELATED APPLICATIONS

This application is the National Stage of International Application No.PCT/EP2004/005362, filed May 18, 2004.

The present invention relates to novel nutraceutical compositionscomprising, as the active ingredients, resveratrol, a derivative,metabolite or analogue thereof, and at least one additional componentselected from EGCG, genistein, vitamin E, polyunsaturated fatty acids,gamma-linolenic acid and vitamin K.

The term “nutraceutical” as used herein denotes a usefulness in both thenutritional and pharmaceutical field of application. Thus, the novelnutraceutical compositions can find use as supplement to food andbeverages, and as pharmaceutical formulations for enteral or parenteralapplication which may be solid formulations such as capsules or tablets,or liquid formulations, such as solutions or suspensions. As will beevident from the foregoing, the term nutraceutical composition alsocomprises food and beverages containing the above-specified activeingredients.

The term “resveratrol, a derivative, metabolite or analogue thereof” asused herein comprises

compounds encompassed by the general formula

wherein A denotes a carbon-carbon double bond which may be trans or cis,and R1, R2, R3, R4, R5 and R6, independently denote hydrogen, hydroxy oretherified or esterified hydroxy groups. While the carbon-carbon doublebond denoted by the symbol A may be trans or cis, formula I above isunderstood to also include cis/trans mixtures. However, compounds offormula I wherein A is a trans carbon-carbon bond are preferred.

Etherified or esterified hydroxy groups maybe derived from unsubstitutedor substituted, straight or branched chain alkyl groups having 1 to 26carbon atoms or from unsubstituted or substituted, straight or branchedchain aliphatic, araliphatic or aromatic carboxylic acids having 1 to 26carbon atoms. Etherified hydroxy groups may further be glycoside groupsand esterified hydroxy groups may further be glucuronide or sulfategroups. Examples of compounds of formula I are resveratrol (R1, R3 andR5=hydrogen, R2, R4 and R6=hydroxy); piceatannol (R3 and R5=hydrogen,R1, R2, R4 and R6=hydroxy), and rhapontigenin (R5=hydrogen, R1, R3, R4and R6=hydroxy, and R2=methoxy). Of primary interest for the purposes ofthe invention is (trans)-resveratrol.

The term “EGCG” as used herein comprises (−)-Epigallocatechin gallateand/or one or more derivatives (esterified forms, glycosides, sulphates)thereof.

The term “polyunsaturated fatty acids” as used herein (also referred toherein as PUFA) denotes a polyunsaturated fatty acid in an esterified(e.g., as triglycerides or ethyl esters) or a free form, particularly anomega-3 polyunsaturated fatty acid such as eicosapentaenoic acid(5,8,11,14,17-eicosapentaenoic acid, EPA) and docosahexaenoic acid(4,7,10,13,16,19-docosahexaenoic acid, DHA), or anomega-6-polyunsaturated fatty acid such as γ-linolenic acid(6,9,12-octadecatrienoic, GLA).

The term “vitamin K” as used herein comprises phylloquinone (vitamin K₁)and naphthoquinol and naphthoquinones having physiological propertiessimilar to vitamin K₁, particularly vitamin K₂ and K₃.

The term “vitamin E” as used herein is a group of compounds based on6-chromanol and includes racemic vitamin E (D,L-α-tocopherol) or naturalvitamin E, as well as derivatives thereof which have biological vitaminE activity, e.g. other tocopherols or tocotrienols or carboxylic acidesters, such as vitamin E acetate, propionate, butyrate or succinate.

The term “genistein” as used herein comprises the aglycone (4′,5,7-trihydroxyisoflavone) and derivatives thereof, e.g., genisteinglycosides, genistein sulfates, genistein glucuronides.

The active ingredients of the composition of this invention havedifferent mechanism of action thus providing synergistic effects in thetreatment or prevention of the diseases specified above, particularlyinflammatory diseases.

Inflammatory disease are one of the most important health problems inthe world. Inflammation is in general a localized protective response ofthe body tissues to invasion of the host by foreign material orinjurious stimuli. The causes of inflammation are infectious agents suchas bacteria, viruses, and parasites; physical agents such as burns orradiation; chemicals like toxins, drugs or industrial agents; andimmunologic reactions such as allergies and autoimmune responses.

Two mains classes of drugs, the cortiocosteroid and the nonsteroidalanti-inflammatory drugs (NSAIDs) are used to treat inflammatorydisorders. NSAIDs and corticosteroids provide essentially symptomaticrelief. Corticosteroids use has declined due to a growing concern aboutthe serious side effects of prolonged use. NSAIDs are among the mostwidely used drugs, primarily for the treatment of pain and inflammation,especially arthritis. However, chronic use of these drugs is alsolimited by their severe side-effects like serious gastrointestinalcomplications, renal toxicity or asthmatic reactions. Thus, there is aneed for new anti-inflammatory agents with weak or no side effects. Inview of the importance of inflammatory disease, in particular rheumaticdisorders, there is a need for anti-inflammatory phytochemicals suitablefor the treatment of non-acute rheumatoid symptoms. Patients withinflammatory diseases have a special interest in treatment considered as“natural” with mild anti-inflammatory effects and without major sideeffects, which can be used for disease prevention and as adjuvanttreatment.

Inflammation is characterized by pain, redness, swelling, heat, andeventual loss of function of the affected area. These symptoms are theresults of a complex series of interactions taking place between thecells of the immune system. The response of the cells results in aninteracting network of three main families of inflammatory mediators:Proteins (e.g.,cytokines, enzymes (e.g., proteases, peroxydase), majorbasic protein (MPC), adhesion molecules (ICAM, VCAM), lipid mediators(e.g., eicosanoids, prostaglandins, leukotrienes, platelet activatingfactor (PAF)), reactive oxygen species (e.g.,hydroperoxides, superoxydeanion O₂ ⁻, nitric oxide (NO). However, many of those mediators ofinflammation are also regulators of normal cellular activity. Thus,deficiencies of inflammatory reactions lead to a compromised host (i.e.infection) while excessive inflammation leads to inflammatory diseasesmediated in part by the over production of several of the abovementioned mediators. The treatment used need to maintain the equilibriumbetween excessive and insufficient inflammatory reaction.

Combination of natural products such as resveratrol and EGCG, genistein,vitamin E, omega-3 PUFA (e.g.,eicosapentaenoic acid, docosahexaenoicacid), borage oil (gamma-linolenic acid), vitamin K with mildanti-inflammatory activity and having different mechanism of action arevery useful for maintaining such a balance.

Acute and chronic inflammation resulting form an excessive biosynthesisof inflammatory mediators is involved in numerous diseases such asarthritis (e.g., osteoarthritis, rheumatoid arthritis), asthma,inflammatory bowel diseases, inflammatory disease of the skin(e.g.,psoriasis, atopic dermatitis) and other chronic diseases with aninflammatory component such as atherosclerosis, heart diseases,diabetes, metabolic syndrome X, cancer, Alzheimer's disease andpre-stages thereof such as mild cognitive impairment.

Rheumatoid arthritis is a chronic inflammatory diseases of the joints.It is characterized by inflammation of the synovium and infiltration ofthe joints by neutrophils, macrophages and lymphocytes. As the diseaseprogresses inflammation leads to synovial proliferation with damage tothe articular cartilage and the bone underlying the cartilage withsubsequent joint deformation and loss of function. Asthma and rheumatoidarthritis are characterised at the molecular level by chronicallyunbalanced expression of cytokines, chemokines, kinins and theirreceptors, adhesion molecules, and inflammatory enzymes such asinducible nitric oxide synthase (iNOS) and the inducible cyclooxygenase(COX-2). Proinflammatory cytokines play also a pivotal role in thedevelopment of osteoarthritis. Psoriasis is one of the most common skinproblem, affecting 1-3% of the human population. Inflammatory boweldisease is a general term used to describe disease such as ulcerativecolitis and Crohn's disease.

Atherosclerosis is currently considered as an inflammatory diseases ofthe vessel wall rather than simply a process of intravascular lipiddeposition. Atherosclerosis results from vascular injury followed by aninflammation. Activated macrophages, T-lymphocytes, and mast cells arepresent in atherosclerotic plaques. Monocyte and lymphocyte activationleads to the release of eicosanoids and cytokines which are implicatedin endothelial damage, as well as in the formation and eventuallyrupture of the atherosclerotic plaques. Finally, circulatinginflammatory markers such as C-reactive protein (CRP), fibrinogen, andinterleukins are increased in groups at high-risk of coronary arterydiseases. Several clinical trials indicate that elevated CRPconcentration correlates with increased risk of coronary, and vascular,events. Thus inflammation appears to play an important role in theinitiation and progression of atheroma formation. Moreover, type 2diabetes and obesity are risk factors for the development of coronaryartery disease and atherosclerosis. Those conditions are associated withinsulin resistance, oxidative stress, and inflammation.

A number of studies in animal models and humans indicate thatinflammation, insulin insensitivity and disturbances in lipid metabolismare linked. Inflammatory mediators, such as plasma interleukin 6 (IL-6),tumor necrosis factor alpha (TNF-α) and CRP are elevated in type 2diabetes and obesity. Adipose tissue can synthesize cytokines such asTNF-α and IL-6 which may promote inflammation in obesity. Therefore,anti-inflammatory agents may play a role not only in the prevention andtreatment of atherosclerosis but also in diabetes and obesity.

Epidemiological studies showed a significant reduction in the risk ofcolorectal, gastric, esophageal, and breast cancers among people whotake non-steroidal anti-inflammatory drugs (NSAIDs) compared with thosenot taking NSAIDs. In animal models NSAIDs significantly reduced tumordevelopment. Increased levels of prostaglandins have been found incancers of breast, colon, lung and pancreas in humans. Moreover, COX-2,is also overexpressed in a variety of tumors. Thus, COX inhibitors mightbe used in cancer prevention and treatment.

Inflammatory events are also associated with the pathophysiology ofAlzheimer's disease. There is evidence of inflammation in the brain ofpatients with Alzheimer's disease. It is characterized by increasedlevels of cytokines and activated microglial cells. Epidemiologicalstudies have suggested that patients taking NSAIDs have a lower risk ofdeveloping Alzheimer's disease than those not taking NSAIDs. Aprotective effect of NSAIDs suggests that the cyclooxygenases might beinvolved in the neurodegenerative process. Therefore, suppression ofexcessive production of inflammatory mediators may prevent and/or slowthe progression of Alzheimer's disease.

Thus, inflammation is not only involved in the classical inflammatorydiseases (e.g.,arthritis, asthma, bowel diseases) but is also associatedwith many chronic diseases (e.g.,atherosclerosis, heart diseases,diabetes, metabolic syndrome X, cancer, Alzheimer disease).

The compositions comprising a combination of active ingredients, i.e.resveratrol or derivatives and at least one additional componentselected from EGCG, genistein, vitamin E, omega-3 PUFA (eicosapentaenoicacid, docosahexaenoic acid), borage oil (gamma-linolenic acid), andvitamin K are particularly useful for the treatment or prevention ofdisease with an inflammatory component. Moreover, a multi-vitamin andmineral supplement maybe added to the nutraceutical compositions of thepresent invention to obtain an adequate amount of an essential nutrient,which is missing in some diets. The multi-vitamin and mineral supplementmay also be useful for disease prevention and protection againstnutritional losses and deficiencies due to lifestyle patterns.

The nutraceutical compositions of the present invention containresveratrol in an amount sufficient to administer to a human adult(weighing about 70 kg) a dosage from about 0.5 mg/day to about 2000mg/day, preferably from about 5 mg/day to about 500 mg/day. Thus, if thenutraceutical composition is a food or beverage the amount of aresveratrol contained therein is suitably in the range from about 0.2 mgto about 500 mg per serving. If the nutraceutical composition is apharmaceutical formulation such formulation may contain from about 0.5mg to about 500 mg per solid dosage unit, e.g., per capsule or tablet,or from about 0.5 mg per daily dose to about 2000 mg per daily dose of aliquid formulation.

EGCG is preferably used in a concentration so that the daily consumptionby a human adult (weighing about 70 kg) is in the range of from 10mg/day to 2000 mg/day. A food or beverage suitably contains about 2 mgto about 500 mg of EGCG per serving. If the nutraceutical composition isa pharmaceutical formulation such formulation may contain a EGCG in anamount from about 5 mg to about 500 mg per dosage unit, e.g., percapsule or tablet, or from about 10 mg per daily dose to about 2000 mgper daily dose of a liquid formulation.

PUFA's are preferably used in a concentration so that the dailyconsumption by a human adult (weighing about 70 kg) is in the range offrom 10 mg/day to 4000 mg/day. A food or beverage suitably containsabout 5 mg to about 1000 mg of a PUFA per serving. If the nutraceuticalcomposition is a pharmaceutical formulation such formulation may containa PUFA in an amount from about 10 mg to about 1000 mg per dosage unit,e.g., per capsule or tablet, or from about 10 mg per daily dose to about4000 mg per daily dose of a liquid formulation.

Genistein is preferably used in a concentration so that the dailyconsumption by a human adult (weighing about 70 kg) is in the range offrom 0.5 mg/day to 2000 mg/day. A food or beverage suitably containsabout 0.2 mg to about 500 mg of genistein per serving. If thenutraceutical composition is a pharmaceutical formulation suchformulation may contain a genistein in an amount from about 0.5 mg toabout 500 mg per dosage unit, e.g., per capsule or tablet, or from about0.5 mg per daily dose to about 2000 mg per daily dose of a liquidformulation.

Vitamin E or its derivative is preferably used in a concentration sothat the daily consumption by a human adult (weighing about 70 kg) is inthe range of from 5 mg/day to 2000 mg/day. A food or beverage suitablycontains about 2 mg to about 500 mg of vitamin E per serving. If thenutraceutical composition is a pharmaceutical formulation suchformulation may contain vitamin E in an amount from about 5 mg to about1000 mg per dosage unit, e.g., per capsule or tablet, or from about 5 mgper daily dose to about 2000 mg per daily dose of a liquid formulation.

Vitamin K is preferably used in a concentration so that the dailyconsumption by a human adult (weighing about 70 kg) is in the range offrom 10 μg/day to 50 mg/day. A food or beverage suitably contains about2 μg per serving to about 20 mg of vitamin K per serving. If thenutraceutical composition is a pharmaceutical formulation suchformulation may contain a vitamin K in an amount from about 10 μg toabout 25 mg per dosage unit, e.g., per capsule or tablet, or from about10 μg per daily dose to about 50 mg per daily dose of a liquidformulation.

The term “serving” as used herein denotes an amount of food or beveragenormally ingested by a human adult with a meal at a time and may range,e.g., from about 100 g to about 500 g.

In one aspect the present invention the compositions may be used as anutritional supplement, e.g., as an additive to a multi-vitaminpreparations comprising vitamins and minerals which are essential forthe maintenance of normal metabolic function but are not synthesized inthe body, especially for the treatment or prevention of inflammatorydiseases.

According to another aspect of the invention the compositions maybepharmaceutical compositions, preferably for enteral application, whichmay be solid or liquid galenical formulation. Examples of solidgalenical formulations are tablets, capsules (e.g. hard or soft shellgelatin capsules), pills, sachets, powders, granules and the like whichcontain the active ingredient together with conventional galenicalcarriers. Any conventional carrier material can be utilized. The carriermaterial can be organic or inorganic inert carrier material suitable fororal administration. Suitable carriers include water, gelatin, gumarabic, lactose, starch, magnesium stearate, talc, vegetable oils, andthe like. Additionally, additives such as flavouring agents,preservatives, stabilizers, emulsifying agents, buffers and the likemaybe added in accordance with accepted practices of pharmaceuticalcompounding. While the individual active ingredients are suitablyadministered in a single composition they may also be administered inindividual dosage units.

Specific combinations of active ingredients in the compositions of thepresent invention comprise

-   Resveratrol and EGCG;-   Resveratrol and vitamin E;-   Resveratrol and PUFA (EPA; DHA; GLA);-   Resveratrol, vitamin E, EGCG;-   Resveratrol, vitamin K, EGCG;-   Resveratrol, vitamin E, PUFA (EPA; DHA; GLA);-   Resveratrol, EGCG, vitamin E and PUFA (EPA; DHA; GLA);-   Resveratrol, vitamin E, PUFA (EPA; DHA; GLA), genistein,-   Resveratrol, EGCG, vitamin E, genistein, PUFA (EPA; DHA; GLA);

Most preferred are the combinations of resveratrol, EGCG, PUFA (EPA;DHA; GLA) with or without vitamin E; and resveratrol, EGCG, vitamin E,genistein, PUFA (EPA; DHA; GLA).

In order to determine anti-inflammatory properties of compounds orcombinations thereof, appropriate cells or cell lines (i.e. whole blood,macrophages, leukocytes) will be activated with inflammatory stimuli invitro in the presence of the compounds. This leads to the secretion ofprostaglandins (i.e. the product of cyclooxygenase-2) and nitric oxide(synthesized by inducible nitric oxide synthase). Due to theiranti-inflammatory effects, compounds will reduce the level of the twometabolites. Similarly, the expression of genes of inflammatory pathwayswill be monitored by quantitative PCR or by micro-array analysis.Anti-inflammatory compounds reduce their expression levels. Additiveand/or synergistic effects of compounds will be identified both at thelevel of specific inflammatory parameters and more generally in the geneexpression profile related to the cellular inflammatory response.

The antiinflammatory effect of a combined therapy with resveratrol andEGCG or EPA can be demonstrated in stimulated macrophages by determiningthe inhibition of the synthesis of nitric oxide and/or PGE₂.

In order to induce an in vitro ‘inflammatory response’, murinemacrophages RAW264.7 were seeded into microtiter plates or 12- wellplates and stimulated with lipopolysaccharide (LPS) without or withgraded amounts of the test substances. Vehicle concentrations (i.e.DMSO) were kept constant. Culture supernatants were harvested afterappropriate periods of time (4 to 24 hours). Prostaglandin E₂ (PGE₂) andnitrite produced from nitric oxide (NO) secreted into the culture mediumwere quantified by ELISA and the Griess reaction, respectively. Thepercentage of inhibition of PGE₂ or NO production at a givenconcentration of the test substances (compared to maximal production byLPS-stimulated cells) was calculated.

The results are indicated in Tables 1 and 2 below:

TABLE 1 Effects of Resveratrol and EGCG on the production of NO and PGE₂in LPS-stimulated macrophages % inhibition NO PGE₂ Resveratrol (10 μM)8.3 26.8 EGCG (10 μM) 4.4 1.8 Resveratrol (10 μM) + EGCG (10 μM)(calculated 12.7 28.6 additive inhibition) Resveratrol (10 μM) + EGCG(10 μM) (actually 42.7 57.0 determined inhibition

TABLE 2 Effects of Resveratrol and EPA on the NO production in LPSstimulated macrophages % inhibition NO Resveratrol (2 μM) 10.0 EPA (12.5μM) 12.9 Resveratrol (2 μM) + EPA (12.5 μM) (calculated additive 22.9inhibition) Resveratrol (2 μM) + EPA (12.5 μM) (actually determined 34.1inhibition

The combined treatment with resveratrol and EGCG exerted a synergisticeffect on NO and PGE₂ production in macrophages (Table 1). Thecombination of resveratrol and eicosapentaenoic Acid (EPA) had asynergistic effect on the production of NO by stimulated macrophages(Table 2). Thus, the data of Tables 1 and 2 show that combinations ofresveratrol and EGCG and resveratrol and EPA exert a synergistic effecton the attenuation of the inflammatory response.

The following Examples illustrate the invention further.

A. Pharmaceutical Compositions May be Prepared by ConventionalFormulation Procedures Using the Ingredients Specified Below:

EXAMPLE 1

Soft Gelatin Capsule

Soft gelatin capsules are prepared by conventional procedures usingingredients specified below:

-   Active ingredients: Resveratrol 10 mg, EPA 200 mg, vitamin E 50 mg-   Other ingredients: glycerol, water, gelatine, vegetable oil

EXAMPLE 2

Hard Gelatin Capsule

Hard gelatin capsules are prepared by conventional procedures usingingredients specified below.

-   Active ingredients: resveratrol 10 mg, EGCG 100 mg, genistein, 5 mg,    vitamin E 50 mg, vitamin K 1 mg-   Other Ingredients:-   Fillers: lactose or cellulose or cellulose derivatives q.s-   Lubricant: magnesium sterate if necessary (0.5%)

EXAMPLE 3

Tablet

Tablets are prepared by conventional procedures using ingredientsspecified below:

-   Active ingredients: resveratrol 5 mg, EGCG 50 mg, vitamin E 20 mg-   Other ingredients: microcrystalline cellulose, silicone dioxide    (SiO2), magnesium stearate, crosscarmellose sodium.

B. Food Items May be Prepared by Conventional Procedures UsingIngredients Specified Below.

EXAMPLE 4

Soft Drink with 30% Juice

-   Active Ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-200 mg/per serving-   EGCG: 2-200 mg/ per serving-   PUFA (EPA; DHA, GLA): 5-500 mg/per serving-   Genistein: 0.2-50 mg/ per serving-   Vitamin E: 5-100 mg/ per serving-   Vitamin K: 0.01-5 mg/ per serving-   Typical serving: 240 ml    I. A Soft Drink Compound is Prepared from the Following Ingredients:

Juice concentrates and water soluble flavours [g] Orange concentrate60.3° Brix, 5.15% acidity 657.99 Lemon concentrate 43.5° Brix, 32.7%acidity 95.96 Orange flavour, water soluble 13.43 Apricot flavour, watersoluble 6.71 Water 26.46 1.2 Color β-Carotene 10% CWS 0.89 Water 67.651.3 Acid and Antioxidant Ascorbic acid 4.11 Citric acid anhydrous 0.69Water 43.18 1.4 Stabilizers Pectin 0.20 Sodium benzoate 2.74 Water 65.601.5 Oil soluble flavours Orange flavour, oil soluble 0.34 Orange oildistilled 0.341.6 Active Ingredients

Active ingredients (this means the active ingredient mentioned above:resveratrol and one or more of the following EGCG, PUFA (EPA; DHA; GLA),genistein, vitamin E and vitamin K) in the concentrations mentionedabove

Fruit juice concentrates and water soluble flavours are mixed withoutincorporation of air. The color is dissolved in deionized water.Ascorbic acid and citric acid is dissolved in water. Sodium benozoate isdissolved in water. The pectin is added unter stirring and dissolvedwhile boiling. The solution is cooled down. Orange oil and oil solubleflavours are premixed. The active ingredients as mentioned under 1.6 aredry mixed and then stirred preferably into the fruit juice concentratemixture (1.1).

In order to prepare the soft drink compound all parts 3.1.1 to 3.1.6 aremixed together before homogenising using a Turrax and then ahigh-pressure homogenizer (p₁=200 bar, P₂=50 bar).

II. A Bottling Syrup is Prepared from the Following Ingredients:

[g] Softdrink compound 74.50 Water 50.00 Sugar syrup 60° Brix 150.00

The ingredients of the bottling syrup are mixed together. The bottlingsyrup is diluted with water to 1 l of ready to drink beverage.

Variations:

Instead of using sodium benzoate, the beverage may be pasteurised. Thebeverage may also be carbonised.

EXAMPLE 5

5 Cereal Bread

-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 50 g

[%] 5 cereal flour 56.8 Water 39.8 Yeast 2.3 Salt 1.1

The yeast is dissolved in a part of the water. All ingredients are mixedtogether to form a dough. Salt is added at the end of the kneading time.After fermentation, the dough is reworked and divided before a loaf isformed. Before baking, the surface of the loaf is brushed with water andsprinkled with flour.

-   Parameters:

Kneading: Spiral kneading system 4 min 1^(st) gear 5 min 2^(nd) gearDough proofing: 60 min Dough temperature: 22-24° C. Proofing time: 30min Baking: Oven: Dutch type oven Baking temperature: 250/220° C. Bakingtime: 50-60 min

EXAMPLE 6

Cookies Type Milano

-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/ per serving-   Typical serving: 30 g

[g] Wheat Flour, type 550 41.0 Sugar 20.5 Fat/Butter 20.5 Whole egg(liquid) 18.0 Lemon Flavour q.s. Baking agent q.s.

All ingredients are added slowly under mixing to form a sweet shortpastry.

Afterwards, the pastry is kept cool (4° C.) for at least 2 hours beforeflattening the pastry to a thickness of approx. 5 mm. Pieces are cut outand brushed with egg yolk on the surface before baking.

Baking:

Oven: fan oven Baking temperature: 180° C. Baking time: 15 min

EXAMPLE 7

Toast

-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 100 g

[%] Wheat Flour, type 550 55.4 Water 33.2 Yeast 2.8 Salt 1.1 Fat/Butter5.5 Malt 0.6 Emulsifier baking agent 1.4

The yeast is dissolved in a part of the water. All ingredients are mixedtogether to form a dough. Salt is added at the end of the kneading time.Afterwards, the dough is reworked, divided and placed in a baking tinfor fermentation. After baking, the loaf is unmoulded directly.

Parameters:

Kneading: Spiral kneading system 5-6 min 1^(st) gear 3-4 min 2^(nd) gearDough proofing: none Dough temperature: 22-24° C. Proofing time: 40 minBaking: Oven: Dutch type oven Baking temperature: 220° C. Baking time:35-40 min

EXAMPLE 8

Yoghurt—Set Type

-   3.5% fat-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 225 g

[%] Full fat milk (3.8% fat) 90.5 Skimmed milk powder 2.0 Sugar 5.0Culture 2.5

The milk is heated to 35° C. before addition of milk powder, stabiliser,sugar and active ingredients. This mixture is heated to 65° C. todissolve all ingredients. Then the mixture is homogenized in ahigh-pressure homogenizer (p₁=150 bar, p₂=50 bar) at 65° C. Thisemulsion is then pasteurised at 80° C. for 20 minutes. After cooling to45° C. natural yoghurt/culture is added and mixed. Then this mixture isfilled into cups and fermented at 45° C. for 3-4 hours until a pH of 4.3is reached and then stored at 4° C.

EXAMPLE 9

Yoghurt—Stirred Type

-   3.5% fat-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 225 g

[%] Full fat milk (3.8% fat) 90.2 Skimmed milk powder 2.0 Stabiliser 0.3Sugar 5.0 Culture 2.5

The milk is heated to 35° C. before addition of milk powder, stabiliser,sugar and active ingredients. This mixture is heated to 65° C. todissolve all ingredients before homogenisation in a high-pressurehomogenizer (p₁=150 bar, p₂=50 bar) at 65° C. This emulsion is thenpasteurised at 80° C. for 20 minutes. After cooling to 45° C. naturalyoghurt/culture is added and mixed, followed by fermentation at 45° C.for 3-4 hours until a pH of 4.3 is reached. After cooling and stirringvigorously, the yoghurt is filled in cups and stored at 4° C.

EXAMPLE 10

Ice Cream

-   8% fat-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-100 mg/per serving-   EGCG: 2-100 mg/per serving-   PUFA (EPA; DHA, GLA): 5-200 mg/per serving-   Genistein: 0.2-20 mg/per serving-   Vitamin E: 5-100 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 85 g

[g] Milk (3.7% fat) 600.00 Cream (35% fat) 166.00 Skim milk powder 49.10Sugar 109.00 Glucose syrup 80% 70.00 Ice cream stabiliser 5.00 Flavorq.s. Color q.s

Sugar, skim milk powder and stabiliser are added to the milk and cream,mixed and heated to 45° C. Then the colour as stock solution and theglucose syrup is added as well as the active ingredients. The mix isheated up and pasteurized (20 min, 80° C.). Then a homogenization steptakes place. Afterwards the mix is cooled down under constant stirringand the flavour is added at 5° C. The mix maturated at 5° C. during atleast 4 h and then passed through an the ice cream machine (overrun ca.100%). The ice cream is filled into cups and stored at −20 to −30° C.

EXAMPLE 11

Wine Gums

-   Active ingredients:-   Resveratrol and one or more additional components selected from    EGCG, PUFA (EPA; DHA; GLA), genistein, vitamin E and vitamin K are    incorporated in this food item-   Resveratrol: 0.2-50 mg/per serving-   EGCG: 2-50 mg/per serving-   PUFA (EPA; DHA, GLA): 5-100 mg/per serving-   Genistein: 0.2-10 mg/per serving-   Vitamin E: 5-10 mg/per serving-   Vitamin K: 0.01-5 mg/per serving-   Typical serving: 30 g

[g] Gelatine 200 Bloom 80.0 Water I 125.0 Sugar crys. 290.0 Water II120.0 Glucose-syrup DE 38 390.0 Citric acid 10.0 Flavour 2.0 Colour q.s.Yield ca 1000.0

Disperse gelatine in water I, stir and dissolve by heating over a streambath or using a microwave. Mix sugar with water II and bring to boilinguntil a clear solution is obtained. Remove from heat source. Mix withglucose syrup while dissolved sugar solution is still hot. Slowly addthe gelatine solution. Let rest until foam on surface can be removed and60-65° C. is reached. Add flavour, citric acid and the colour solutionas well as active ingredients under stirring. Deposit into mouldsprinted into starch trays and let sit for at least 48 hours at RT.Remove starch powder and polish with oil or wax. Dry at RT and packageinto airtight pouch.

1. A method for treating or reducing inflammation in a subject in needof such treatment which comprises administering to the subject acompound of formula I, in combination with (−)-epigallocatechin gallate(EGCG), wherein the compound of formula I is:

wherein A is a carbon-carbon double bond, and R1, R2, R3, R4, R5 and R6are independently selected from hydrogen, hydroxy, etherified hydroxygroups and esterified hydroxy groups; a cis or trans isomer thereof, ora mixture of cis and trans isomers thereof.
 2. The method of claim 1wherein the compound of formula I is resveratrol.
 3. The method of claim2 wherein the resveratrol is administered in an amount of from about 0.5mg/day to about 2000 mg/day and wherein the EGCG is in an amount of fromabout 10 mg/day to about 2000 mg/day.
 4. The method of claim 2 whereinthe resveratrol and the EGCG are administered in a unit dosage form offrom about 0.5 mg to about 500 mg of resveratrol and from about 5 mg toabout 500 mg EGCG.